Strategy of nitrogen acquisition and utilization by carnivorous Dionaea muscipula

Plant carnivory represents an exceptional means to acquire N. Snap traps of Dionaea muscipula serve two functions, and provide both N and photosynthate. Using ¹³C/¹⁵N-labelled insect powder, we performed feeding experiments with Dionaea plants that differed in physiological state and N status (spring vs. autumn plants). We measured the effects of ¹⁵N uptake on light-saturated photosynthesis (A _max), dark respiration (R _D) and growth. Depending on N status, insect capture briefly altered the dynamics of R _D/A _max, reflecting high energy demand during insect digestion and nutrient uptake, followed by enhanced photosynthesis and growth. Organic N acquired from insect prey was immediately redistributed, in order to support swift renewal of traps and thereby enhance probability of prey capture. Respiratory costs associated with permanent maintenance of the photosynthetic machinery were thereby minimized. Dionaea’s strategy of N utilization is commensurate with the random capture of large prey, occasionally transferring a high load of organic nutrients to the plant. Our results suggest that physiological adaptations to unpredictable resource availability are essential for Dionaea’s success with regards to a carnivorous life style.     

The influence of secretion elicitors and external pH on the kinetics of D-alanine uptake by the trap lobes of Dionaea muscipula Ellis (Venus's Flytrap)

Simple kinetic techniques were used to examine the mechanism of D-alanine uptake by the adaxial surfaces of the trap lobes of Dionaea muscipula Ellis (Venus's Flytrap.) On the basis of these analyses, the uptake of D-alanine was found to depend on the time during which the trap lobes were inoculated with elicitors of secretion before excision and measurement of uptake. Disks taken from traps that had not been subjected to a preceding period of inoculation with secretion elicitors showed a low basal rate of uptake which was neither pH-dependent nor exhibited saturation with respect to external D-alanine concentration. Disks from preinoculated traps, on the other hand, displayed an enhanced rate of uptake which showed both pH-dependence and saturation with respect to external D-alanine concentration. The capacity for enhanced uptake was lost upon prolonged inoculation or when inoculation was stopped. Of the compounds tested, only elicitors of secretion caused an enhancement of uptake. The enhanced rate of D-alanine uptake is temperature-sensitive with a Q₁₀ characteristic of a mediated process. Uncouplers cause an instantaneous abolition of uptake whereas the effects of terminal-oxidase inhibitors are time-dependent. The pH-dependence of uptake is inferred to result from an increased affinity of the carrier system for D-alanine at low pH values. Although the ionic state of D-alanine is relatively unaffected over the pH range examined, a decrease in the external pH from 6.0 to 3.8 decreases the apparent K _m for uptake by four-fold but increases V _max by only 30%. It is concluded that the acid secreted by the digestive glands of Dionaea plays a direct role in facilitating the uptake of amino acids from the trap cavity.     

Accurate determination of mean cell volume by isotope dilution in erythrocyte populations with variable deformability.

Variations in erythrocyte deformability and morphology lead to artifacts in electronic determinations of mean cellular volume (MCV) by the aperture-impedance method. The micropipette-aspiration technique loses accuracy when applied to severely aberrant cells such as dense sickle cells. A new light-scattering technique requires that the cells be capable of undergoing isovolumetric sphering. In contrast, the isotope-dilution (ID) method measures absolute mean volume and is free of artifacts associated with abnormal deformability or morphology. It does not depend on any algorithms or correction factors and does not subject the cells to any stringent processing, not even centrifugation. The ID method can be used to determine the mean volume of red cells in hypo- or hypertonic media or in the presence of pharmacologic agents. It requires no more than a 1-ml aliquot of suspended cells at a hematocrit of at least 30%. The cells can be readily recovered, washed, and reused. Using EDTA labeled with 57Co as an extracellular space marker we have used ID to determine the MCV of fractionated normal human red blood cells (RBC), unfractionated RBC containing SS hemoglobin, and RBC from four other mammalian species. In the case of human RBC obtained from eight normal donors, we obtained mean MCV values (+/- SD) of 83.6 +/- 3.0, 87.5 +/- 3.9, and 76.5 +/- 5.3 fl for unfractionated and top and bottom 10% density fractions, respectively. The value 83.6 is significantly lower than the generally accepted range of 89-91 indicated by electronic analyzers calibrated against spun microhematocrits. The discrepancy of about 7% can account for the difference between mean cell hemoglobin concentration (MCHC) data determined by a calibrated Coulter Counter and corresponding data obtained with paired samples using a cyanmethemoglobin procedure specified in NCCLS Standard H15-A and corrected for trapped plasma.     

Comparison of predator–prey interactions with and without intraguild predation by manipulation of the nitrogen source

Theory predicts that intraguild predation leads to different community dynamics than the trophic cascades of a linear food chain. However, experimental comparisons of these two food‐web modules are rare. Mixotrophic plankton species combine photoautotrophic and heterotrophic nutrition by grazing upon other phytoplankton species. We found that the mixotrophic chrysophyte Ochromonas can grow autotrophically on ammonium, but not on nitrate. This offered a unique opportunity to compare predator–prey interactions in the presence and absence of intraguild predation, without changing the species composition of the community. With ammonium as nitrogen source, Ochromonas can compete with its autotrophic prey for nitrogen and therefore acts as intraguild predator. With nitrate, Ochromonas acts solely as predator, and is not in competition with its prey for nitrogen. We parameterized a simple intraguild predation model based on chemostat experiments with monocultures of Ochromonas and the toxic cyanobacterium Microcystis. Subsequently, we tested the model predictions by inoculating Ochromonas into the Microcystis monocultures, and vice versa. The results showed that Microcystis was a better competitor for ammonium than Ochromonas. In agreement with theoretical predictions, Microcystis was much more strongly suppressed by intraguild predation on ammonium than by top–down predation on nitrate. Yet, Microcystis persisted at very low population densities, because the type III functional response of Ochromonas implied that the grazing pressure upon Microcystis became low when Microcystis was rare. Our results provide experimental support for intraguild predation theory, and indicate that intraguild predation may enable biological control of microbial pest species.     

氮高效利用基因型水稻生育后期氮素分配与转运特性

选择前期筛选出的氮高效利用基因型水稻为试验材料,以低效利用基因型为对照,采用土培试验,在低氮(100 mg·kg^-1)和正常施氮(200 mg·kg^-1)下,研究了高效和低效基因型水稻生育后期不同器官的氮素分配量、转运量和转运效率差异.结果表明: 与低效基因型水稻相比,高效基因型在低氮条件下,仍能保持较高的产量和氮素利用效率,其产量为低效基因型的1.75倍,氮肥利用率高达50.9%,而低效基因型仅为36.4%.与正常施氮相比,低氮更有利于提高氮素在高效基因型穗部的分配量,穗部积累量在扬花期、灌浆期和成熟期分别增加了34.2%、2.5%和0.5%,而低效基因型在灌浆期和成熟期却分别降低了23.5%和15.6%.不同施氮水平下,氮素在高效基因型不同器官的分配比例为扬花期:叶＞茎鞘＞根＞穗,灌浆期:穗＞叶＞茎鞘＞根,成熟期:穗＞茎鞘＞叶＞根,随着生育期的推进,穗部的分配比例明显增加.在低氮和正常施氮下,高效基因型氮素转运量表现为叶＞茎鞘＞根,而低效基因型表现为茎鞘＞叶＞根;高效基因型氮素转运效率分别为60.8%、60.3%,分别为低效基因型的1.67、155倍.因此,高效基因型抽穗后叶片较高的转运效率为籽粒的灌浆结实奠定了良好基础.     

Impacts of a tertiary treated municipal wastewater effluent on the carbon and nitrogen stable isotope signatures of two darter species (Etheostoma blennioides and E. caeruleum) in a small receiving environment

This study contrasted the influence of tertiary treated municipal wastewater effluent (MWWE) exposure on the abundance, food selection (stomach contents), and carbon (δ¹³C) and nitrogen (δ¹⁵N) stable isotope signatures of Rainbow Darter (Etheostoma caeruleum) and Greenside Darter (Etheostoma blennioides) in a small receiving stream in southern Ontario, Canada. Despite tertiary treatment resulting in relatively high effluent quality, there is continued concern for environmental degradation downstream of the effluent outfall because of the relatively small size of the receiving environment and recent studies that have indicated changes occurring in relative fish abundance and stable isotope signatures. In July and August of 2009 fish and benthic invertebrate communities were examined along with analysis of δ¹³C and δ¹⁵N of the most common species to determine the effects of effluent exposure on the food web. Rainbow Darter increased in abundance and their δ¹⁵N values were enriched immediately downstream of the effluent outfall throughout the summer (July and August). In contrast, while δ¹⁵N of Greenside Darter followed a similar pattern in July, they were not enriched in August. The benthic community was changed immediately downstream of the outfall and the δ¹⁵N of invertebrates was enriched, similar to that observed in Rainbow Darter. Stomach contents of the two darter species diverged the most in the summer possibly explaining some of the changes in δ¹³C and δ¹⁵N among sites. It is possible that Greenside Darter fed on less enriched food downstream of the outfall, or fed outside of the effluent plume during the summer. Rainbow Darter may be able to exploit the changes in habitat and prey composition, resulting in differences in relative abundance of darters immediately downstream of the effluent outfall.     

Mechanism of the reaction catalyzed by dihydrofolate reductase from Escherichia coli: pH and deuterium isotope effects with NADPH as the variable substrate

The variations with pH of the kinetic parameters and primary deuterium isotope effects for the reaction of NADPH with dihydrofolate reductase from Escherichia coli have been determined. The aims of the investigations were to elucidate the chemical mechanism of the reaction and to obtain information about the location of the rate-limiting steps. The V and V/KNADPH profiles indicate that a single ionizing group at the active center of the enzyme must be protonated for catalysis, whereas the Ki profiles show that the binding of NADPH to the free enzyme and of ATP-ribose to the enzyme-dihydrofolate complex is pH independent. From the results of deuterium isotope effects on V/KNADPH, it is concluded that NADPH behaves as a sticky substrate. It is this stickiness that raises artificially the intrinsic pK value of 6.4 for the Asp-27 residue of the enzyme-dihydrofolate complex [Howell, E. E., Villafranca, J. E., Warren, M. S., Oatley, S. J., & Kraut, J. (1986) Science (Washington, D.C.) 231, 1123] to an observed value of 8.9. Thus, the binary enzyme complex is largely protonated at neutral pH. The elevation of the intrinsic pK value of 6.4 for the ternary enzyme-NADPH-dihydrofolate complex to 8.5 is not due to the kinetic effects of substrates. Rather, it is the consequence of the lower, pH-independent rate of product release and the faster pH-dependent catalytic step. At neutral pH, the proportion of enzyme present as a protonated ternary enzyme-substrate complex is sufficient to keep catalysis faster than product release.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparative utilization of inorganic and organic compounds as sole nitrogen sources by the submergent duckweed,Lemna trisulca L.

The capability of the submergentLemna trisulca L. to utilize various inorganic and organic sources of nitrogen was studied using both non-axenic and axenic cultures. When doubling time for frond production was measured, the nitrogen sources in order of effectiveness were urea, aspartic acid, nitrate, glutamic acid, arginine, ammonium and casein hydrolysate. Nitrite supported a relatively rapid growth rate after an initial lag of 7 days. Other parameters of growth such as fresh or dry weight per frond or ohlorophyll content did not oorrelate well with rate of frond production. Casein hydrolysate and urea were found to elicit a morphology different from that seen in cultures containing the other nitrogen sources. These preferences for source of nitrogen were different than those known for the emergent species ofLemnaceae. The unique value ofL. trisulca as a subject for plant physiological research is discussed. This study also provides a possible explanation for the existence of nutritional niches existing in aquatio ecosystems containing several different species ofLemnaceae.     

Longitudinal pattern of resource utilization by aquatic consumers along a disturbed subtropical urban river: Estimating the relative contribution of resources with stable isotope analysis

The utilization of food resources by aquatic consumers reflects the structure and functioning of river food webs. In lotic water systems, where food availability and predator–prey relationships vary with gradient changes in physical conditions, understanding diet assimilation by local communities is important for ecosystem conservation. In the subtropical Liuxi River, southern China, the relative contribution of basal resources to the diet assimilation of functional feeding groups (FFGs) was determined by stable carbon (¹³C) and nitrogen (¹⁵N) isotope analyses. The output of Bayesian mixing models showed that diatom‐dominated periphyton (epilithic biofilm), aquatic C₃ plants (submerged hydrophytes), and suspended particulate organic matter (SPOM) associated with terrestrial C₃ plants contributed the most to the diet assimilation of FFGs in the upper, middle, and lower reaches, respectively. The relative contribution of consumer diet assimilation was weighted by the biomass (wet weight, g/m²) of each FFG to reflect resource utilization at the assemblage level. From the upper to the lower reaches, the spatial variation in the diet assimilation of fish and invertebrate assemblages could be summarized as a longitudinal decrease in periphyton (from 57%–76% to <3%) and an increase in SPOM (from <7% to 51%–68%) with a notable midstream increase in aquatic C₃ plants (23%–48%). These results indicate that instream consumers in the Liuxi River rely more on autochthonous production (e.g., periphyton and submerged hydrophytes) than on terrestrially derived allochthonous matter (e.g., terrestrial plants). The pattern of resource utilization by consumers in the mid‐upper Liuxi River is consistent with findings from other open subtropical and neotropical rivers and provides evidence for the riverine productivity model. Our study indicates that protecting inherent producers in rivers (e.g., periphyton and submerged hydrophytes) and restoring their associated habitats (e.g., riffles with cobble substrate) are conducive to aquatic ecosystem management.     

Determination of the mechanism of human malic enzyme with natural and alternate dinucleotides by isotope effects.

Human malic enzyme was studied by steady state kinetics, deuterium isotope effects, and 13C isotope effects with both the physiological dinucleotide cofactor and several alternate cofactors. The log V vs pH profile with NAD revealed two pK(a) values too close to be separately determined, but with an average value of 7.33. The log V/K vs pH profile with NAD revealed two pK(a) values at 7.4 and 5.6. Deuterium and 13C isotope effects indicate that the mechanism of human malic enzyme is stepwise with both NAD and epsilonNAD, but that hyperconjugation in the transition state for hydride transfer is detectable only with the former. With thioNAD and APAD, the isotope effects do not clearly indicate whether the mechanism is stepwise or concerted. The intrinsic 13C isotope effect for decarboxylation was calculated to be 1.0485 by measurement of the partition ratio of oxaloacetate in the presence of NADH and human malic enzyme (decarboxylation to pyruvate/reduction to malate = 2.33). The isotope effect and partitioning data suggest that the energy barrier for decarboxylation of oxaloacetate is not as high relative to the barrier for reduction of oxaloacetate as with the chicken liver enzyme.     

Expression and secretion of the bovine coronavirus hemagglutinin-esterase glycoprotein by insect cells infected with recombinant baculoviruses.

A cDNA fragment representing the hemagglutinin-esterase (HE) gene of bovine coronavirus (BCV) was inserted into the genome of Autographa californica nuclear polyhedrosis virus. Infection of insect cells with the recombinant virus resulted in the production of a 120-kilodalton disulfide-linked dimeric form of the BCV HE polypeptide. Deletion of the carboxy-terminal hydrophobic domain from the HE polypeptide resulted in secretion of a dimeric form of the truncated HE polypeptide. The acetylesterase activity of the BCV HE was detectable in insect cells expressing the BCV hemagglutinin and was inhibited by two monoclonal antibodies which also inhibit hemagglutination.     

Methods of improving the utilization of cereal straw by ruminants. III. A note on the effect of ensiling straw treated with urea

Straw treated with urea was offered to lambs either freshly prepared (EN₀) or after ensiling with urea for 30 days (EN₃₀). The lambs also received 10 g dry matter (DM) of shredded, artificially dried grass per day per kg of metabolic liveweight. Ensiling straw significantly increased its intake from 32 to 41 g DM per day per kg of metabolic liveweight, and its DM digestibility from 375 to 450 g per kg.     

Studies on the purification of wastes from the nitrogen fertilizer industry by intensive algal cultures. IV. growth of Chlorella vulgaris in wastes with high nitrogen content in continuous and intermittent light.

The growth and utilization of nitrogen by intensive Chlorella vulgaris in wastes from production of urea, containing 1300 mg NH4+-N and 4000 mg urea-N/1, was investigated. In these conditions only Chlorella vulgaris AA strain, adapted to high concentrations of ammonia nitrogen, was able to grow. The elimination of nitrogen by continuous cultures was 750 mg urea-N/1 with 5-day flow rate. A considerable part of the urea was hydrolized by urease bacteria and removed in the form of NH3. The effect of intermittent light on the growth of algae was also studied. The better growth than in continuous light, was obtained with alternate one hour periods of light and darkness. Good results were also obtained with the use of 12 hour light and 12 hour darkness.     

Stimulatory effect of cold adaptation on glucose utilization by brown adipose tissue. Relationship with changes in the glucose transporter system

The effect of cold adaptation (4 degrees C) on the in vivo glucose utilization and on the number and properties of the glucose transporters has been studied in brown adipose tissue of normal rats. Glucose utilization was assessed in vivo by the 2-deoxyglucose method. Glucose transporters in plasma and microsomal membranes were quantified by the [3H]cytochalasin B-binding assay. After cold adaptation the in vivo glucose utilization by brown adipose tissue increased 21-fold compared to controls (22 degrees C). The number of glucose transporters in plasma membranes of brown adipose tissue increased from 75 to 436 pmol/g tissue and that of total glucose transporters (plasma + microsomal membranes) from 438 to 754 pmol/g tissue. In addition, cold adaptation increased the Hill coefficient of the plasma membrane transporter for cytochalasin B from 0.90 to 2.03 and decreased the Kd from 100 to 54 nM. This study shows that cold adaptation promotes: a translocation of glucose transporters from an intracellular pool to plasma membranes; an increased number of plasma membrane glucose transporters unaccounted for by the translocation process (e.g. "de novo" synthesis); an increase in the Hill coefficient for cytochalasin B that could also represent changes in the properties of the transporters vis-à-vis glucose, (e.g. positive cooperativity); and a decrease in the Kd value for cytochalasin B.